Jiang Han-Liang, Jiang Li-Ming, Han Wei-Dong
Department of Pulmonology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Zhejiang, China.
J BUON. 2015 Jul-Aug;20(4):1094-100.
In the present study, we have analyzed the regulation of Wnt/β-catenin signaling in lung adenocarcinoma stem cells (CSCs), that are responsible for tumor recurrence.
Lung cancer samples were studied for the presence of cancer stem like cells and analyzed by flow cytometry. Then, the sorted cells were analyzed for the stem cell surface markers and Wnt/β-catenin signaling pathways. Moreover, the sorted side population (SP) and non-SP cells were also subjected to drug resistance assay.
Western blot analysis showed that the protein level of β-catenin was highly upregulated in fluorescence activated cells (FACs) sorted SP cells which led to elevated expression of stem cell protein Oct-4 that is responsible for SP cells' self-renewal. RT-PCR revealed that the relative mRNA expression level of Wnt target gene cyclin D was significantly higher (p<0.01) in SP cells, enhancing thus the cell proliferation rate and clone formation efficiency. In addition, the matrigel invasion assay revealed that SP cells were highly invasive than non-SP cells.
In the present study we demonstrated that lung adenocarcinoma samples contain a small population of tumor-initiating SP cells which possess the characteristic features of CSCs. Wnt/β-catenin mediated increased expression of β-catenin, Oct-4 and cyclin D in SP cells but not in non-SP cells was also observed. FACs-purified SP cells are resistant to a number of chemotherapeutic drugs. Our data suggest that the use of novel anticancer drugs, targeting Wnt/β-catenin signaling pathways, may help eradicate the lung cancers stem cells.
在本研究中，我们分析了肺腺癌干细胞（CSCs）中Wnt/β-连环蛋白信号通路的调控情况，这些干细胞是肿瘤复发的原因。
研究肺癌样本中癌症干细胞样细胞的存在情况，并通过流式细胞术进行分析。然后，对分选的细胞进行干细胞表面标志物和Wnt/β-连环蛋白信号通路分析。此外，分选的侧群（SP）细胞和非SP细胞也进行了耐药性检测。
蛋白质印迹分析表明，在荧光激活细胞分选的SP细胞中，β-连环蛋白的蛋白质水平高度上调，这导致负责SP细胞自我更新的干细胞蛋白Oct-4表达升高。逆转录聚合酶链反应显示，Wnt靶基因细胞周期蛋白D的相对mRNA表达水平在SP细胞中显著更高（p<0.01），从而提高了细胞增殖率和克隆形成效率。此外，基质胶侵袭试验显示，SP细胞比非SP细胞具有更高的侵袭性。
在本研究中，我们证明肺腺癌样本中含有一小群具有CSCs特征的肿瘤起始SP细胞。还观察到Wnt/β-连环蛋白介导SP细胞中β-连环蛋白、Oct-4和细胞周期蛋白D的表达增加，而非SP细胞中未增加。流式细胞术纯化的SP细胞对多种化疗药物耐药。我们的数据表明，使用针对Wnt/β-连环蛋白信号通路的新型抗癌药物可能有助于根除肺癌干细胞。
